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The article provides information on challenges to the implementation of social programs for safety of workplace in hospitals including lack of customer centric approach, violence among safety professionals, and weak resolutions.

Purpose Dental professionals are exposed to hazardous noise levels on a daily basis in clinical practice. The purpose of this study was to compare the hearing status of dental hygienists who utilize ultrasonic scalers in the workplace compared to age-matched control participants (non-dental hygienists) who were not exposed to ultrasonic noise. Methods A convenience sample of nineteen dental hygienists (experimental) and nineteen non-dental hygienists (control) was recruited for this study. A matched pairs design was utilized; participants in each group were matched based on age and gender to eliminate confounding variables. The testing procedure consisted of an audiologist performing a series of auditory tests including otoacoustic emissions test, pure-tone audiometry, and tympanometry on the experimental and control groups. Results In the right ear, there were notable differences from 1000 Hz - 10,000 Hz and in the left ear from 6000 Hz - 10,000 Hz, with higher hearing thresholds in the experimental group of dental hygienists. While 56% of the univariate tests conducted on how many days were worked per week showed statistical significance, the regression line slope indicated those that worked more days had better hearing statuses. The variables for years in practice for dental hygienists, how many of those years were full-time employment, and how many years the dental hygienist had used an ultrasonic scaling device, also had many significant univariate tests for the experimental group only. These variables were more likely to serve as proxies representing true noise exposure. The paired t-test between the groups demonstrated statistically significant differences between the experimental and control group at 9000 Hz in both ears. Conclusion While results from this study demonstrated various qualitative differences in hearing status of the control group (non-dental hygienists) and experimental group (dental hygienists), age was found to be the most critical variable. Furthermore, this data demonstrated differences in hearing status based on various frequencies between dental hygienists and age-matched controls that should be further explored with a larger population., (Copyright © 2024 The American Dental Hygienists’ Association.)

Tertiary Lymphoid Structures (TLS) are lymphoid structures commonly associated with improved survival of cancer patients and response to immunotherapies. However, conflicting reports underscore the need to consider TLS heterogeneity and multiple features such as TLS size, composition, and maturation status, when assessing their functional impact. With the aim of gaining insights into TLS biology and evaluating the prognostic impact of TLS maturity in Non-Small Cell Lung Carcinoma (NSCLC), we developed a multiplex immunofluorescent (mIF) panel including T cell (CD3, CD8), B cell (CD20), Follicular Dendritic cell (FDC) (CD21, CD23) and mature dendritic cell (DC-LAMP) markers. We deployed this panel across a cohort of primary tumor resections from NSCLC patients (N=406) and established a mIF image analysis workstream to specifically detect TLS structures and evaluate the density of each cell phenotype. We assessed the prognostic significance of TLS size, number, and composition, to develop a TLS scoring system representative of TLS biology within a tumor. TLS relative area, (total TLS area divided by the total tumor area), was the most prognostic TLS feature (C-index: 0.54, p = 0.04). CD21 positivity was a marker driving the favorable prognostic impact, where CD21 + CD23 - B cells (C-index: 0.57, p = 0.04) and CD21 + CD23 - FDC (C-index: 0.58, p = 0.01) were the only prognostic cell phenotypes in TLS. Combining the three most robust prognostic TLS features: TLS relative area, the density of B cells, and FDC CD21 + CD23 - we generated a TLS scoring system that demonstrated strong prognostic value in NSCLC when considering the effect of age, sex, histology, and smoking status. This TLS Score also demonstrated significant association with Immunoscore, EGFR mutational status and gene expression-based B-cell and TLS signature scores. It was not correlated with PD-L1 status in tumor cells or immune cells. In conclusion, we generated a prognostic TLS Score representative of the TLS heterogeneity and maturity undergoing within NSCLC tissues. This score could be used as a tool to explore how TLS presence and maturity impact the organization of the tumor microenvironment and support the discovery of spatial biomarker surrogates of TLS maturity, that could be used in the clinic., Competing Interests: Authors JB, HA, PP, EJ, SW, JB, MSa, FN and MSu were employed by the company AstraZeneca and are shareholders. Authors FS, EJ, MV, TG, AA, HH and MH-B were employed by the company AstraZeneca. Author JG was employed by the company Veracyte. The remaining author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The authors declare that this study received funding from AstraZeneca. Additionally, Author TG received research funding from Idera Pharmaceuticals, consulting fees from Mendus, LAVA Therapeutics and GE Health, and is shareholder of LAVA Therapeutics. Author RV received research funding from Genmab BV. However, none of these funders, except AstraZeneca, were involved in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision., (Copyright © 2024 Berthe, Poudel, Segerer, Jennings, Ng, Surace, Andoni, Testori, Saraiya, Vuko, Hessel, Heininen-Brown, Blando, Jones, Willis, Galon, van de Ven, de Gruijl and Angell.)

Purpose: To assess the efficacy of various machine learning (ML) algorithms in predicting late-stage colorectal cancer (CRC) diagnoses against the backdrop of socio-economic and regional healthcare disparities., Methods: An innovative theoretical framework was developed to integrate individual- and census tract-level social determinants of health (SDOH) with sociodemographic factors. A comparative analysis of the ML models was conducted using key performance metrics such as AUC-ROC to evaluate their predictive accuracy. Spatio-temporal analysis was used to identify disparities in late-stage CRC diagnosis probabilities., Results: Gradient boosting emerged as the superior model, with the top predictors for late-stage CRC diagnosis being anatomic site, year of diagnosis, age, proximity to superfund sites, and primary payer. Spatio-temporal clusters highlighted geographic areas with a statistically significant high probability of late-stage diagnoses, emphasizing the need for targeted healthcare interventions., Conclusions: This research underlines the potential of ML in enhancing the prognostic predictions in oncology, particularly in CRC. The gradient boosting model, with its robust performance, holds promise for deployment in healthcare systems to aid early detection and formulate localized cancer prevention strategies. The study's methodology demonstrates a significant step toward utilizing AI in public health to mitigate disparities and improve cancer care outcomes.

Neoadjuvant chemoimmunotherapy improves pathologic complete response rate and event-free survival in patients with resectable non-small cell lung cancer (NSCLC) versus chemotherapy alone. NeoCOAST was the first randomized, multidrug platform trial to examine novel neoadjuvant immuno-oncology combinations for patients with resectable NSCLC, using major pathologic response (MPR) rate as the primary endpoint. Eighty-three patients received a single cycle of treatment: 26 received durvalumab (anti-PD-L1) monotherapy, 21 received durvalumab plus oleclumab (anti-CD73), 20 received durvalumab plus monalizumab (anti-NKG2A), and 16 received durvalumab plus danvatirsen (anti-STAT3 antisense oligonucleotide). MPR rates were higher for patients in the combination arms versus durvalumab alone. Safety profiles for the combinations were similar to those of durvalumab alone. Multiplatform immune profiling suggested that improved MPR rates in the durvalumab plus oleclumab and durvalumab plus monalizumab arms were associated with enhanced effector immune infiltration of tumors, interferon responses and markers of tertiary lymphoid structure formation, and systemic functional immune cell activation., Significance: A neoadjuvant platform trial can rapidly generate clinical and translational data using candidate surrogate endpoints like MPR. In NeoCOAST, patients with resectable NSCLC had improved MPR rates after durvalumab plus oleclumab or monalizumab versus durvalumab alone and tumoral transcriptomic signatures indicative of augmented immune cell activation and function. See related commentary by Cooper and Yu, p. 2306. This article is featured in Selected Articles from This Issue, p. 2293., (©2023 The Authors; Published by the American Association for Cancer Research.)

Purpose: MEDI0680 is a humanized anti-programmed cell death-1 (PD-1) antibody, and durvalumab is an anti-PD-L1 antibody. Combining treatment using these antibodies may improve efficacy versus blockade of PD-1 alone. This phase II study evaluated antitumor activity and safety of MEDI0680 plus durvalumab versus nivolumab monotherapy in immunotherapy-naïve patients with advanced clear-cell renal cell carcinoma who received at least one prior line of antiangiogenic therapy., Patients and Methods: Patients received either MEDI0680 (20 mg/kg) with durvalumab (750 mg) or nivolumab (240 mg), all intravenous, every 2 weeks. The primary endpoint was investigator-assessed objective response rate (ORR). Secondary endpoints included best overall response, progression-free survival (PFS), safety, overall survival (OS), and immunogenicity. Exploratory endpoints included changes in circulating tumor DNA (ctDNA), baseline tumor mutational burden, and tumor-infiltrated immune cell profiles., Results: Sixty-three patients were randomized (combination, n = 42; nivolumab, n = 21). ORR was 16.7% [7/42; 95% confidence interval (CI), 7.0-31.4] with combination treatment and 23.8% (5/21; 95% CI, 8.2-47.2) with nivolumab. Median PFS was 3.6 months in both arms; median OS was not reached in either arm. Because of adverse events, 23.8% of patients discontinued MEDI0680 and durvalumab and 14.3% of patients discontinued nivolumab. In the combination arm, reduction in ctDNA fraction was associated with longer PFS. ctDNA mutational analysis did not demonstrate an association with response in either arm. Tumor-infiltrated immune profiles showed an association between immune cell activation and response in the combination arm., Conclusions: MEDI0680 combined with durvalumab was safe and tolerable; however, it did not improve efficacy versus nivolumab monotherapy., (©2022 The Authors; Published by the American Association for Cancer Research.)

Wildfires have increased in frequency and magnitude and pose a significant public health challenge. The principal objective of this study was to assess the impact of wildfire smoke on respiratory peak flow performance of patients exposed to two different wildfire events. This longitudinal study utilized an observational approach and a cohort study design with a patient-level clinical dataset from a local outpatient allergy clinic ( n = 842). Meteorological data from a local weather station served as a proxy for smoke exposure because air quality measurements were not available. This study found that there were decreases in respiratory peak flow among allergy clinic patients one year after each wildfire event. For every one percent increase in wind blowing from the fire towards the community, there was, on average, a 2.21 L per minute decrease in respiratory peak flow. This study observed an effect on respiratory peak flow performance among patients at a local allergy clinic one year after suspected exposure to wildfire smoke. There are likely multiple reasons for the observation of this relationship, including the possibility that wildfire smoke may enhance allergic sensitization to other allergens or that wildfire smoke itself may elicit a delayed immune response.

Rhabdomyosarcoma is the most common malignant soft tissue tumor in pediatric age. It can affect any anatomical location. Alveolar histological subtype usually presents lesions on the extremities in older children. The most common metastatic sites are the lung, bone marrow, bone and lymph node. We describe a case of alveolar rhabdomyosarcoma with cardiac metastasis in a pediatric patient, a rare presentation of the pathology., Competing Interests: None, (Sociedad Argentina de Pediatría.)

Renal medullary carcinoma (RMC) is a highly lethal malignancy that mainly afflicts young individuals of African descent and is resistant to all targeted agents used to treat other renal cell carcinomas. Comprehensive genomic and transcriptomic profiling of untreated primary RMC tissues was performed to elucidate the molecular landscape of these tumors. We found that RMC was characterized by high replication stress and an abundance of focal copy-number alterations associated with activation of the stimulator of the cyclic GMP-AMP synthase interferon genes (cGAS-STING) innate immune pathway. Replication stress conferred a therapeutic vulnerability to drugs targeting DNA-damage repair pathways. Elucidation of these previously unknown RMC hallmarks paves the way to new clinical trials for this rare but highly lethal malignancy., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Background: We sought to ascertain the immune effector function of pembrolizumab within the glioblastoma (GBM) microenvironment during the therapeutic window., Methods: In an open-label, single-center, single-arm phase II "window-of-opportunity" trial in 15 patients with recurrent (operable) GBM receiving up to 2 pembrolizumab doses before surgery and every 3 weeks afterward until disease progression or unacceptable toxicities occurred, immune responses were evaluated within the tumor., Results: No treatment-related deaths occurred. Overall median follow-up time was 50 months. Of 14 patients monitored, 10 had progressive disease, 3 had a partial response, and 1 had stable disease. Median progression-free survival (PFS) was 4.5 months (95% CI: 2.27, 6.83), and the 6-month PFS rate was 40%. Median overall survival (OS) was 20 months, with an estimated 1-year OS rate of 63%. GBM patients' recurrent tumors contained few T cells that demonstrated a paucity of immune activation markers, but the tumor microenvironment was markedly enriched for CD68+ macrophages., Conclusions: Immune analyses indicated that pembrolizumab anti-programmed cell death 1 (PD-1) monotherapy alone can't induce effector immunologic response in most GBM patients, probably owing to a scarcity of T cells within the tumor microenvironment and a CD68+ macrophage preponderance., (© The Author(s) 2019. Published by Oxford University Press on behalf of the Society for Neuro-Oncology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Tumors with high mutational burden (TMB) tend to be responsive to immune checkpoint blockade (ICB) because there are neoantigens available for targeting by reinvigorated T cells, whereas those with low TMB demonstrate limited clinical responses. To determine whether antigen-specific T cell responses can be elicited after treatment with ICB in cancers that have a low TMB, we conducted a clinical trial with ipilimumab in 30 patients with metastatic castration-resistant prostate cancer. We identified two distinct cohorts by survival and progression times: "favorable" ( n = 9) and "unfavorable" ( n = 10). Patients in the favorable cohort had high intratumoral CD8 T cell density and IFN-γ response gene signature and/or antigen-specific T cell responses. Two patients with a relatively low TMB had T cell responses against unique neoantigens. Moreover, six of nine patients in the favorable group are still alive at the time of analysis, with survival ranging from 33 to 54 months after treatment. All 10 patients in the unfavorable cohort have succumbed to their disease and had survival ranging from 0.6 to 10.3 months. Collectively, our data indicate that immunological correlates associated with effector T cell responses are observed in patients with metastatic prostate cancer who benefit from ICB., (Copyright © 2020 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Treatment with immune checkpoint blockade (ICB) has revolutionized cancer therapy. Until now, predictive biomarkers 1-10 and strategies to augment clinical response have largely focused on the T cell compartment. However, other immune subsets may also contribute to anti-tumour immunity 11-15 , although these have been less well-studied in ICB treatment 16 . A previously conducted neoadjuvant ICB trial in patients with melanoma showed via targeted expression profiling 17 that B cell signatures were enriched in the tumours of patients who respond to treatment versus non-responding patients. To build on this, here we performed bulk RNA sequencing and found that B cell markers were the most differentially expressed genes in the tumours of responders versus non-responders. Our findings were corroborated using a computational method (MCP-counter 18 ) to estimate the immune and stromal composition in this and two other ICB-treated cohorts (patients with melanoma and renal cell carcinoma). Histological evaluation highlighted the localization of B cells within tertiary lymphoid structures. We assessed the potential functional contributions of B cells via bulk and single-cell RNA sequencing, which demonstrate clonal expansion and unique functional states of B cells in responders. Mass cytometry showed that switched memory B cells were enriched in the tumours of responders. Together, these data provide insights into the potential role of B cells and tertiary lymphoid structures in the response to ICB treatment, with implications for the development of biomarkers and therapeutic targets.

Immune checkpoint therapy with anti-CTLA-4 and anti-PD-1/PD-L1 has revolutionized the treatment of many solid tumors. However, the clinical efficacy of immune checkpoint therapy is limited to a subset of patients with specific tumor types 1,2 . Multiple clinical trials with combinatorial immune checkpoint strategies are ongoing; however, the mechanistic rationale for tumor-specific targeting of immune checkpoints is elusive. To garner an insight into tumor-specific immunomodulatory targets, we analyzed 94 patients representing five different cancer types, including those that respond relatively well to immune checkpoint therapy and those that do not, such as glioblastoma multiforme, prostate cancer and colorectal cancer. Through mass cytometry and single-cell RNA sequencing, we identified a unique population of CD73 hi macrophages in glioblastoma multiforme that persists after anti-PD-1 treatment. To test if targeting CD73 would be important for a successful combination strategy in glioblastoma multiforme, we performed reverse translational studies using CD73 -/- mice. We found that the absence of CD73 improved survival in a murine model of glioblastoma multiforme treated with anti-CTLA-4 and anti-PD-1. Our data identified CD73 as a specific immunotherapeutic target to improve antitumor immune responses to immune checkpoint therapy in glioblastoma multiforme and demonstrate that comprehensive human and reverse translational studies can be used for rational design of combinatorial immune checkpoint strategies.

Introduction: OX-40 co-stimulatory signaling plays a role in mounting anti-tumor immune responses and clinical trials targeting this pathway are ongoing. However, the association of with OX-40 protein expression with clinical outcomes and pathological features in non-small cell lung cancer (NSCLC) are largely unknown., Methods: Surgically-resected stage I-III NSCLC specimens (N = 100) were stained by immunohistochemistry (IHC) for the following immune markers: OX-40, PD-L1, PD-1, CD3, CD4, CD8, CD45RO, CD57, CD68, FOXP3, granzyme B, and ICOS. Immune-related markers mRNA expression were also assessed. We evaluated the association of OX-40 levels with major clinicopathologic variables, including molecular driver mutations., Results: OX-40 IHC expression was observed in all tested tumors, predominantly localized in the membrane of the tumor immune infiltrate, and was not associated with a specific clinicopathologic or molecular subtype. High OX-40 expression levels measured by IHC median score were associated with better overall survival (OS) (p = 0.002), independent of CD3/CD8, PD-L1, and ICOS expression. High OX-40 IHC score was associated with increased expression of immune-related genes such as CD3, IFN-gamma, ICOS, CD8, CXCL9, CXCL10, CCL5, granzyme K., Conclusions: High OX-40 IHC expression in the tumor immune infiltrate is associated with favorable prognosis and increased levels of immune-related genes including IFN-gamma in patients with surgically resected stage I-III NSCLC. Its prognostic utility is independent of PD-L1 and other common markers of immune activation. High OX-40 expression potentially identifies a unique subgroup of NSCLC that may benefit from co-stimulation with OX-40 agonist antibodies and potentially enhance the efficacy of existing immune checkpoint therapies.

Purpose: Determine the differential effect of a FcγR-binding, mIgG2a anti-GITR antibody in mouse tumor models, and characterize the tumor microenvironment for the frequency of GITR expression in T-cell subsets from seven different human solid tumors. Experimental Design: For mouse experiments, wild-type C57BL/6 mice were subcutaneously injected with MC38 cells or B16 cells, and BALB/c mice were injected with CT26 cells. Mice were treated with the anti-mouse GITR agonist antibody 21B6, and tumor burden and survival were monitored. GITR expression was evaluated at the single-cell level using flow cytometry (FC). A total of 213 samples were evaluated for GITR expression by IHC, 63 by FC, and 170 by both in seven human solid tumors: advanced hepatocellular carcinoma, non-small cell lung cancer (NSCLC), renal cell carcinoma, pancreatic carcinoma, head and neck carcinoma, melanoma, and ovarian carcinoma., Results: The therapeutic benefit of 21B6 was greatest in CT26 followed by MC38, and was least in the B16 tumor model. The frequency of CD8 T cells and effector CD4 T cells within the immune infiltrate correlated with response to treatment with GITR antibody. Analysis of clinical tumor samples showed that NSCLC, renal cell carcinoma, and melanoma had the highest proportions of GITR-expressing cells and highest per-cell density of GITR expression on CD4 + Foxp3 + T regulatory cells. IHC and FC data showed similar trends with a good correlation between both techniques., Conclusions: Human tumor data suggest that NSCLC, renal cell carcinoma, and melanoma should be the tumor subtypes prioritized for anti-GITR therapy development., (©2019 American Association for Cancer Research.)

Adoptive transfer of tumor-reactive T cells (ACT) has led to modest clinical benefit in the treatment of solid tumors. Failures with this therapy are primarily due to inadequate infiltration and poor function of adoptively transferred cells in the tumor microenvironment. To improve the efficacy of ACT, we combined ACT with dual blockade of CTLA-4 and PD-1. Treatment with anti-CTLA-4 plus anti-PD-1 compared with monotherapy resulted in durable antitumor responses, enhanced effector function of ACT, utilizing PMEL-1 transgenic (Tg + ) CD8 + T cells, and improved survival. Using PMEL-1ICOS -/- mice, we showed that deletion of the inducible T-cell costimulator (ICOS) receptor abolished the therapeutic benefits, with selective downregulation of Eomesodermin (Eomes), interferon gamma (IFNγ), and perforin. Higher expression of IFNγ and Eomes was noted in human ICOS hi CD8 + T cells compared with ICOS low counterparts. Together, our data provide direct evidence that ACT combined with immune-checkpoint therapy confers durable antitumor responses, which largely depended on CD8 + T-cell-intrinsic expression of ICOS. Our study provides a foundation of testing combinatorial therapy of ACT of CD8 T cells and dual blocking of CTLA-4 and PD-1 in patients with melanoma., (©2019 American Association for Cancer Research.)

Immune checkpoint (IC) therapy provides substantial benefits to cancer patients but can also cause distinctive toxicities termed immune-related adverse events (irAEs). Biomarkers to predict toxicities will be necessary to improve management of patients receiving IC therapy. We relied on serological analysis of recombinant cDNA expression libraries to evaluate plasma samples from patients treated with IC therapy and identified autoantibodies, both in pretreatment and on-treatment samples prior to the development of irAEs, which correlate with the development of immune-related hypophysitis (anti-GNAL and anti-ITM2B autoantibodies) and pneumonitis (anti-CD74 autoantibody). We developed an enzyme-linked immunosorbent assay and tested additional patient samples to confirm our initial findings. Collectively, our data suggest that autoantibodies may correlate with irAEs related to IC therapy, and specific autoantibodies may be detected early for the management of irAEs., Competing Interests: Competing interest statement: J.P.A. has ownership in Jounce, Neon, BioAtla, Forty-Seven, Apricity, Polaris, Marker Therapeutics, Codiak, ImaginAb, Hummingbird, Dragonfly, Lytix, and Tvardi Therapeutics and serves as a consultant for Jounce, Kite Pharma, Neon, Amgen, Forty-Seven, Apricity, Polaris, Marker Therapeutics, Codiak, ImaginAb, Tvardi Therapeutics, Lytix, Hummingbird, and Dragonfly. P.S. has ownership in Jounce, Neon, Constellation, Oncolytics, BioAtla, Forty-Seven, Apricity, Polaris, Marker Therapeutics, Codiak, ImaginAb, Lytix, Hummingbird, and Dragonfly and serves as a consultant for Constellation, Jounce, Kite Pharma, Neon, BioAtla, Pieris Pharmaceuticals, Oncolytics Biotech, Merck, BioMx, Forty-Seven, Polaris, Apricity, Marker Therapeutics, Codiak, ImaginAb, Hummingbird, Lytix, and Dragonfly. J.G. serves as a consultant for ARMO Biosciences, AstraZeneca, CRISPR Therapeutics, Jounce, Nektar, Polaris, Pfizer, and Symphogen. C.G.D. and J.P.A. are coauthors on a 2016 workshop report.

In hepatocellular carcinoma (HCC), surgical resection is associated with high recurrence rate, and no effective adjuvant therapy currently exists. We initiated a pilot randomized trial of perioperative immunotherapy with nivolumab and ipilimumab for resectable HCC. Here, we provide an illustrative report of a case that achieved a complete response and report immunologic correlates of this complete pathologic response to perioperative immunotherapy. Clinical response was correlated with an increase in CD8 + T-cell infiltration, with an increase in two effector T-cell clusters. This study is ongoing, and the final results may contribute to a paradigm shift in the perioperative treatment of HCC, leading to the incorporation of immunotherapy in the curative setting., (©2019 American Association for Cancer Research.)

Background: Phenotypic characterization of immune cells in the bone marrow (BM) of patients with acute myeloid leukemia (AML) is lacking., Methods: T-cell infiltration was quantified on BM biopsies from 13 patients with AML, and flow cytometry was performed on BM aspirates (BMAs) from 107 patients with AML who received treatment at The University of Texas MD Anderson Cancer Center. The authors evaluated the expression of inhibitory receptors (programmed cell death protein 1 [PD1], cytotoxic T-lymphocyte antigen 4 [CTLA4], lymphocyte-activation gene 3 [LAG3], T-cell immunoglobulin and mucin-domain containing-3 [TIM3]) and stimulatory receptors (glucocorticoid-induced tumor necrosis factor receptor-related protein [GITR], OX40, 41BB [a type 2 transmembrane glycoprotein receptor], inducible T-cell costimulatory [ICOS]) on T-cell subsets and the expression of their ligands (41BBL, B7-1, B7-2, ICOSL, PD-L1, PD-L2, and OX40L) on AML blasts. Expression of these markers was correlated with patient age, karyotype, baseline next-generation sequencing for 28 myeloid-associated genes (including P53), and DNA methylation proteins (DNA methyltransferase 3α, isocitrate dehydrogenase 1[IDH1], IDH2, Tet methylcytosine dioxygenase 2 [TET2], and Fms-related tyrosine kinase 3 [FLT3])., Results: On histochemistry evaluation, the T-cell population in BM appeared to be preserved in patients who had AML compared with healthy donors. The proportion of T-regulatory cells (Tregs) in BMAs was higher in patients with AML than in healthy donors. PD1-positive/OX40-positive T cells were more frequent in AML BMAs, and a higher frequency of PD1-positive/cluster of differentiation 8 (CD8)-positive T cells coexpressed TIM3 or LAG3. PD1-positive/CD8-positive T cells were more frequent in BMAs from patients who had multiply relapsed AML than in BMAs from those who had first relapsed or newly diagnosed AML. Blasts in BMAs from patients who had TP53-mutated AML were more frequently positive for PD-L1., Conclusions: The preserved T-cell population, the increased frequency of regulatory T cells, and the expression of targetable immune receptors in AML BMAs suggest a role for T-cell-harnessing therapies in AML., (© 2018 The Authors. Cancer published by Wiley Periodicals, Inc. on behalf of American Cancer Society.)

Preclinical models have shown that blocking PD-1/PD-L1 pathways enhances antileukemic responses. Azacitidine upregulates PD-1 and IFNγ signaling. We therefore conducted this single-arm trial, in which patients with relapsed/refractory (R/R) acute myeloid leukemia (AML) were treated with azacitidine 75 mg/m 2 days 1 to 7 intravenously or subcutaneously with nivolumab 3 mg/kg intravenously on days 1 and 14, every 4 to 6 weeks. For the seventy patients who were treated, the median age was 70 years (range, 22-90) and the median number of prior therapies received was 2 (range, 1-7). The overall response rate (ORR) was 33%, including 15 (22%) complete remission/complete remission with insufficient recovery of counts, 1 partial response, and 7 patients with hematologic improvement maintained >6 months. Six patients (9%) had stable disease >6 months. The ORR was 58% and 22%, in hypomethylating agent (HMA)-naïve ( n = 25) and HMA-pretreated ( n = 45) patients, respectively. Grade 3 to 4 immune-related adverse events occurred in 8 (11%) patients. Pretherapy bone marrow and peripheral blood CD3 and CD8 were significantly predictive for response on flow cytometry. CTLA4 was significantly upregulated on CD4 + Teff in nonresponders after 2 and 4 doses of nivolumab. Azacitidine and nivolumab therapy produced an encouraging response rate and overall survival in patients with R/R AML, particularly in HMA-naïve and salvage 1 patients. Pretherapy bone marrow aspirate and peripheral blood CD3 percentage may be biomarkers for patient selection. SIGNIFICANCE: Azacitidine in combination with nivolumab appeared to be a safe and effective therapy in patients with AML who were salvage 1, prior hypomethylator-naïve, or had increased pretherapy CD3 + bone marrow infiltrate by flow cytometry or IHC. Bone marrow CD3 and CD8 are relatively simple assays that should be incorporated to select patients in future trials. This article is highlighted in the In This Issue feature, p. 305 ., (©2018 American Association for Cancer Research.)

Immune checkpoint therapy (ICT) has transformed cancer treatment in recent years; however, treatment response is not uniform across tumor types. The tumor immune microenvironment plays a critical role in determining response to ICT; therefore, understanding the differential immune infiltration between ICT-sensitive and ICT-resistant tumor types will help to develop effective treatment strategies. We performed a comprehensive analysis of the immune tumor microenvironment of an ICT-sensitive tumor (melanoma, n = 44) and an ICT-resistant tumor (pancreatic cancer, n = 67). We found that a pancreatic tumor has minimal to moderate infiltration of CD3, CD4, and CD8 T cells; however, the immune infiltrates are predominantly present in the stromal area of the tumor and are excluded from tumoral area compared with melanoma, where the immune infiltrates are primarily present in the tumoral area. Metastatic pancreatic ductal adenocarcinomas (PDACs) had a lower infiltration of total T cells compared with resectable primary PDACs, suggesting that metastatic PDACs have poor immunogenicity. Further, a significantly higher number of CD68 + macrophages and VISTA + cells (also known as V-domain immunoglobulin suppressor of T cell activation) were found in the pancreatic stromal area compared with melanoma. We identified VISTA as a potent inhibitory checkpoint that is predominantly expressed on CD68+ macrophages on PDACs. These data suggest that VISTA may be a relevant immunotherapy target for effective treatment of patients with pancreatic cancer., Competing Interests: Conflict of interest statement: J.P.A. is an inventor and recipient of royalties from intellectual property licensed to Bristol-Meyer Squibb, Merck, and Jounce. He is a member of the scientific advisory board for Jounce Therapeutics, Neon Therapeutics, Amgen, Apricity, BioAlta, Forty-Seven, Tvardi Therapeutics, TapImmune, ImaginAB, Codiak Biosciences, and Marker Therapeutics. J.P.A. and P.S. own a patent licensed to Jounce Therapeutics. P.S. serves as a consultant for Constellation, Jounce Therapuetics, Kite Pharma, Neon Therapeutics, BioAtla, Pieris Pharmaceuticals, Oncolytics Biotech, Merck, BioMx, Forty-Seven, Polaris, Apricity, Marker Therapeutics, Codiak, ImaginAB, and TapImmune. She also has stock ownership in Jounce, Neon Therapeutics, Constellation, Oncolytics, BioAtlanta, Forty-Seven, Apricity, Polaris, Marker Therapeutics, Codiak, ImaginAB, and TapImmune. A.S. is an employee of Janssen.

Preclinical studies suggest that treatment with neoadjuvant immune checkpoint blockade is associated with enhanced survival and antigen-specific T cell responses compared with adjuvant treatment 1 ; however, optimal regimens have not been defined. Here we report results from a randomized phase 2 study of neoadjuvant nivolumab versus combined ipilimumab with nivolumab in 23 patients with high-risk resectable melanoma ( NCT02519322 ). RECIST overall response rates (ORR), pathologic complete response rates (pCR), treatment-related adverse events (trAEs) and immune correlates of response were assessed. Treatment with combined ipilimumab and nivolumab yielded high response rates (RECIST ORR 73%, pCR 45%) but substantial toxicity (73% grade 3 trAEs), whereas treatment with nivolumab monotherapy yielded modest responses (ORR 25%, pCR 25%) and low toxicity (8% grade 3 trAEs). Immune correlates of response were identified, demonstrating higher lymphoid infiltrates in responders to both therapies and a more clonal and diverse T cell infiltrate in responders to nivolumab monotherapy. These results describe the feasibility of neoadjuvant immune checkpoint blockade in melanoma and emphasize the need for additional studies to optimize treatment regimens and to validate putative biomarkers.

Clinical responses to immunotherapy have been associated with augmentation of preexisting immune responses, manifested by heightened inflammation in the tumor microenvironment. However, many tumors have a noninflamed microenvironment, and response rates to immunotherapy in melanoma have been <50%. We approached this problem by utilizing immunotherapy (CTLA-4 blockade) combined with chemotherapy to induce local inflammation. In murine models of melanoma and prostate cancer, the combination of chemotherapy and CTLA-4 blockade induced a shift in the cellular composition of the tumor microenvironment, with infiltrating CD8 + and CD4 + T cells increasing the CD8/Foxp3 T-cell ratio. These changes were associated with improved survival of the mice. To translate these findings into a clinical setting, 26 patients with advanced melanoma were treated locally by isolated limb infusion with the nitrogen mustard alkylating agent melphalan followed by systemic administration of CTLA-4 blocking antibody (ipilimumab) in a phase II trial. This combination of local chemotherapy with systemic checkpoint blockade inhibitor resulted in a response rate of 85% at 3 months (62% complete and 23% partial response rate) and a 58% progression-free survival at 1 year. The clinical response was associated with increased T-cell infiltration, similar to that seen in the murine models. Together, our findings suggest that local chemotherapy combined with checkpoint blockade-based immunotherapy results in a durable response to cancer therapy. Cancer Immunol Res; 6(2); 189-200. ©2018 AACR ., (©2018 American Association for Cancer Research.)

Objectives: The aim of this study was to examine tumor-infiltrating lymphocytes (TILs) and their prognostic value in patients with pancreatic ductal adenocarcinoma (PDAC) after neoadjuvant therapy., Methods: Intratumoral CD4, CD8, and FOXP3 lymphocytes were examined by immunohistochemistry using a computer-assisted quantitative analysis in 136 PDAC patients who received neoadjuvant therapy and pancreaticoduodenectomy. The results were correlated with clinicopathological parameters and survival., Results: High CD4 TILs in treated PDAC were associated with high CD8 TILs (P = 0.003), differentiation (P = 0.04), and a lower frequency of recurrence (P = 0.02). Patients with high CD4 TILs had longer disease-free survival and overall survival (OS) than did patients with low CD4 TILs (P < 0.01). The median OS of patients with a high CD8/FOXP3 lymphocyte ratio (39.5 [standard deviation, 6.1] months) was longer than that of patients with a low CD8/FOXP3 lymphocyte ratio (28.3 [standard deviation, 2.3] months; P = 0.01). In multivariate analysis, high CD4 TILs were an independent prognostic factor for disease-free survival (hazard ratio, 0.49; 95% confidence interval, 0.30-0.81; P = 0.005) and OS (hazard ratio, 0.54; 95% confidence interval, 0.33-0.89; P = 0.02)., Conclusions: High level of CD4 lymphocytes is associated with tumor differentiation and lower recurrence and is an independent prognostic factor for survival in PDAC patients treated with neoadjuvant therapy.

Obesity is a prognostic risk factor in the progression of prostate cancer; however, the molecular mechanisms involved are unclear. In this study, we provide preclinical proof of concept for the role of a proinflammatory CXCL12-CXCR4/CXCR7 signaling axis in an obesity-driven mouse model of myc-induced prostate cancer. Analysis of the stromal vascular fraction from periprostatic white adipose tissue from obese HiMyc mice at 6 months of age revealed a dramatic increase in mRNAs encoding various chemokines, cytokines, growth factors, and angiogenesis mediators, with CXCL12 among the most significantly upregulated genes. Immunofluorescence staining of ventral prostate tissue from obese HiMyc mice revealed high levels of CXCL12 in the stromal compartment as well as high staining for CXCR4 and CXCR7 in the epithelial compartment of tumors. Prostate cancer cell lines derived from HiMyc tumors (HMVP2 and derivative cell lines) displayed increased protein expression of both CXCR4 and CXCR7 compared with protein lysates from a nontumorigenic prostate epithelial cell line (NMVP cells). CXCL12 treatment stimulated migration and invasion of HMVP2 cells but not NMVP cells. These effects of CXCL12 on HMVP2 cells were inhibited by the CXCR4 antagonist AMD3100 as well as knockdown of either CXCR4 or CXCR7. CXCL12 treatment also produced rapid activation of STAT3, NFκB, and MAPK signaling in HMVP2 cells, which was again attenuated by either AMD3100 or knockdown of CXCR4 or CXCR7. Collectively, these data suggest that CXCL12 secreted by stromal cells activates invasiveness of prostate cancer cells and may play a role in driving tumor progression in obesity. Targeting the CXCL12-CXCR4/CXCR7 axis could lead to novel approaches for offsetting the effects of obesity on prostate cancer progression. Cancer Res; 77(18); 5158-68. ©2017 AACR ., (©2017 American Association for Cancer Research.)

To date, anti-CTLA-4 (ipilimumab) or anti-PD-1 (nivolumab) monotherapy has not been demonstrated to be of substantial clinical benefit in patients with prostate cancer. To identify additional immune-inhibitory pathways in the prostate-tumor microenvironment, we evaluated untreated and ipilimumab-treated tumors from patients in a presurgical clinical trial. Levels of the PD-L1 and VISTA inhibitory molecules increased on independent subsets of macrophages in treated tumors. Our data suggest that VISTA represents another compensatory inhibitory pathway in prostate tumors after ipilimumab therapy.

Background: Squamous cell carcinoma of the anal canal (SCCA) is a rare malignancy associated with infection by human papillomavirus (HPV). No consensus treatment approach exists for the treatment of metastatic disease. Because intratumoral HPV oncoproteins upregulate immune checkpoint proteins such as PD-1 to evade immune-mediated cytotoxicity, we did a trial of the anti-PD-1 antibody nivolumab for patients with metastatic SCCA., Methods: We did this single-arm, multicentre, phase 2 trial at ten academic centres in the USA. We enrolled patients with treatment-refractory metastatic SCCA, who were given nivolumab every 2 weeks (3 mg/kg). The primary endpoint was response according to Response Evaluation Criteria in Solid Tumors, version 1.1, in the intention-to-treat population. At the time of data cutoff, the study was ongoing, with patients continuing to receive treatment. The study is registered with ClinicalTrials.gov, number NCT02314169., Results: We screened 39 patients, of whom 37 were enrolled and received at least one dose of nivolumab. Among the 37 patients, nine (24% [95% CI 15-33]) had responses. There were two complete responses and seven partial responses. Grade 3 adverse events were anaemia (n=2), fatigue (n=1), rash (n=1), and hypothyroidism (n=1). No serious adverse events were reported., Interpretation: To our knowledge, this is the first completed phase 2 trial of immunotherapy for SCCA. Nivolumab is well tolerated and effective as a monotherapy for patients with metastatic SCCA. Immune checkpoint blockade appears to be a promising approach for patients with this orphan disease., Funding: National Cancer Institute/Cancer Therapy Evaluation Program, the HPV and Anal Cancer Foundation, the E B Anal Cancer Fund, The University of Texas MD Anderson Moon Shots Program, and an anonymous philanthropic donor., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Purpose: We investigated the correlation between immunohistochemical PD-L1 expression and tumor-associated immune cells (TAICs) density in non-small cell lung carcinoma (NSCLC) and correlated them with clinicopathologic variables., Experimental Design: Tumor tissue specimens from 254 stage I-III NSCLCs [146 adenocarcinomas; 108 squamous cell carcinomas (SCCs)] were examined. PD-L1 expression in malignant cells and macrophages and the density of TAICs expressing CD3, CD4, CD8, CD57, granzyme B, CD45RO, PD-1, FOXP3, and CD68 were evaluated using immunohistochemistry and image analysis., Results: Malignant cells PD-L1 H-score > 5 was detected in 23% of adenocarcinomas and 31% of SCCs, and no significant differences were detected comparing both histologies; the median H-score in macrophages was significantly higher in SCC than in adenocarcinoma (P < 0.001). In adenocarcinoma, high malignant cells PD-L1 expression and high TAIC density correlated with solid histology, smoking history, and airflow limitation. Multivariate analysis demonstrated that high CD57-positive cell density correlated with better recurrence-free survival (RFS; P = 0.0236; HR, 0.457) and overall survival (OS; P = 0.0261; HR, 0.481) rates for SCC. High CD68-positive cell density in intratumoral compartment correlated with better RFS (P = 0.0436; HR, 0.553) for adenocarcinoma. The combination of low CD4/CD8/C68-positive cell density and PD-L1 H-score >5 in malignant cells identified small subset of adenocarcinomas with worse outcomes (RFS: P = 0.036; HR, 4.299; OS: P = 0.00034; HR, 5.632)., Conclusions: We detected different PD-L1 expression and TAIC density patterns in NSCLC. Distinct groups of tumor microenvironment correlated with NSCLC clinicopathologic features, including outcome. Clin Cancer Res; 22(24); 6278-89. ©2016 AACR., (©2016 American Association for Cancer Research.)

Unlabelled: Immune checkpoint blockade represents a major breakthrough in cancer therapy; however, responses are not universal. Genomic and immune features in pretreatment tumor biopsies have been reported to correlate with response in patients with melanoma and other cancers, but robust biomarkers have not been identified. We studied a cohort of patients with metastatic melanoma initially treated with cytotoxic T-lymphocyte-associated antigen-4 (CTLA4) blockade (n = 53) followed by programmed death-1 (PD-1) blockade at progression (n = 46), and analyzed immune signatures in longitudinal tissue samples collected at multiple time points during therapy. In this study, we demonstrate that adaptive immune signatures in tumor biopsy samples obtained early during the course of treatment are highly predictive of response to immune checkpoint blockade and also demonstrate differential effects on the tumor microenvironment induced by CTLA4 and PD-1 blockade. Importantly, potential mechanisms of therapeutic resistance to immune checkpoint blockade were also identified., Significance: These studies demonstrate that adaptive immune signatures in early on-treatment tumor biopsies are predictive of response to checkpoint blockade and yield insight into mechanisms of therapeutic resistance. These concepts have far-reaching implications in this age of precision medicine and should be explored in immune checkpoint blockade treatment across cancer types. Cancer Discov; 6(8); 827-37. ©2016 AACR.See related commentary by Teng et al., p. 818This article is highlighted in the In This Issue feature, p. 803., Competing Interests: No other potential conflicts of interest were disclosed., (©2016 American Association for Cancer Research.)

A cell line was established from ventral prostate (VP) tumors of one-year-old Hi-Myc mice. These cells, called HMVP2 cells, are LinnegSca-1highCD49fhigh with high CD44 and CD29 expression and express CK14, Sca-1 and CD49f (but not CK8), suggesting basal-epithelial characteristics. Furthermore, HMVP2 cells form spheroids and both the cells and spheroids produce tumors in syngeneic mice. After four days of culture, HMVP2 spheroids underwent a gradual transition from LinnegSca-1highCD49fhigh expression to LinnegSca-1lowCD49flow while a subpopulation of the cells retained the original LinnegSca-1highCD49fhigh expression pattern. Additional cell subpopulations expressing Lin positive markers were also present suggesting further differentiation of HMVP2 spheroids. Two additional highly tumorigenic cell lines (HMVP2A1 and HMVP2A2) were isolated from HMVP2 cells after subsequent tumor formation in FVB/N mice. Concurrently, we also established cell lines from the VP of 6 months old Hi-Myc mice (named as HMVP1) and FVB/N mice (called NMVP) having less aggressive growth properties compared to the other three cell lines. AR expression was reduced in HMVP2 cells compared to NMVP and HMVP1 cells and almost absent in HMVP2A1 and HMVP2A2 cells. These cell lines will provide valuable tools for further mechanistic studies as well as preclinical studies to evaluate preventive and/or therapeutic agents for prostate cancer., Competing Interests: The authors disclose no potential conflicts of interest.

Unlabelled: The practice of household bush burning in Grenada occurs frequently, though it is not well documented. The effects of the emissions from bush burning on respiratory health of the population have never been researched in Grenada. The goal of the study was to measure the frequency of bush burning and to investigate the relationship between bush burning practice and respiratory health in Grenada. In this cross-sectional study, a questionnaire was used to gather information from households in the parishes of St. George and St. Andrew, Grenada. In total, 225 participants were recruited and their responses on household bush burning and respiratory symptoms were analyzed. Self-reported data showed that the practice of bush burning was conducted by 43% (n = 96) of the participants as a regular practice (every month) and 86% (n = 192) of participants stated that their neighbors burn bush regularly. The most common lower respiratory symptom associated with bush burning was dry cough (31.4%). The participants who engage in the practice of bush burning had a statistically significant, higher prevalence of sinusitis symptoms (OR: 2.1, CI 95%: 1.1-3.9) and had slightly higher prevalence of cough (OR: 1.6, CI 95%: 0.9-2.8). Prevalences of physician-diagnosed asthma and sinusitis were 12.3% and 31.2%, respectively. Conducting studies on the health effects of bush burning in different settings and with different practices, such as the household bush burning in our current study, could help to improve public health in the developing world., Implications: Household disposal of waste is a significant issue in the developing world. In particular, residential bush burning is a common practice in tropical regions. This study demonstrates that the common practice of bush burning in the Caribbean is associated with respiratory symptoms and demonstrates the need for better management of residential yard waste. Burning of yard waste results in potentially significant exposures to air pollution and therefore alternative disposal practices need to be available. There is a need to increase awareness of the importance of avoiding exposure to the air pollutants generated during bush burning among communities in the Caribbean.

In this study, we compared the effect of oral administration of metformin (MET) and rapamycin (RAPA) alone or in combination on prostate cancer development and progression in HiMyc mice. MET (250 mg/kg body weight in the drinking water), RAPA (2.24 mg/kg body weight microencapsulated in the diet), and the combination inhibited progression of prostatic intraepithelial neoplasia lesions to adenocarcinomas in the ventral prostate (VP). RAPA and the combination were more effective than MET at the doses used. Inhibition of prostate cancer progression in HiMyc mice by RAPA was associated with a significant reduction in mTORC1 signaling that was further potentiated by the combination of MET and RAPA. In contrast, treatment with MET alone enhanced AMPK activation, but had little or no effect on mTORC1 signaling pathways in the VP of HiMyc mice. Further analyses revealed a significant effect of all treatments on prostate tissue inflammation as assessed by analysis of the expression of cytokines, the presence of inflammatory cells and NFκB signaling. MET at the dose used appeared to reduce prostate cancer progression primarily by reducing tissue inflammation whereas RAPA and the combination appeared to inhibit prostate cancer progression in this mouse model via the combined effects on both mTORC1 signaling as well as on tissue inflammation. Overall, these data support the hypothesis that blocking mTORC1 signaling and/or tissue inflammation can effectively inhibit prostate cancer progression in a relevant mouse model of human prostate cancer. Furthermore, combinatorial approaches that target both pathways may be highly effective for prevention of prostate cancer progression in men., (©2015 American Association for Cancer Research.)

Effective workplace violence (WPV) prevention programs are essential, yet challenging to implement in healthcare. The aim of this study was to identify major barriers to implementation of effective violence prevention programs. After reviewing the related literature, the authors describe their research methods and analysis and report the following seven themes as major barriers to effective implementation of workplace violence programs: a lack of action despite reporting; varying perceptions of violence; bullying; profit-driven management models; lack of management accountability; a focus on customer service; and weak social service and law enforcement approaches to mentally ill patients. The authors discuss their findings in light of previous studies and experiences and offer suggestions for decreasing WPV in healthcare settings. They conclude that although many of these challenges to effective implementation of workplace violence programs are both within the program itself and relate to broader industry and societal issues, creative innovations can address these issues and improve WPV prevention programs.

Effective workplace violence (WPV) prevention programs are essential, yet challenging to implement in healthcare. The aim of this study was to identify major barriers to implementation of effective violence prevention programs. After reviewing the related literature, the authors describe their research methods and analysis and report the following seven themes as major barriers to effective implementation of workplace violence programs: a lack of action despite reporting; varying perceptions of violence; bullying; profit-driven management models; lack of management accountability; a focus on customer service; and weak social service and law enforcement approaches to mentally ill patients. The authors discuss their findings in light of previous studies and experiences and offer suggestions for decreasing WPV in healthcare settings. They conclude that although many of these challenges to effective implementation of workplace violence programs are both within the program itself and relate to broader industry and societal issues, creative innovations can address these issues and improve WPV prevention programs.

Despite much recent progress, prostate cancer continues to represent a major cause of cancer-related mortality and morbidity in men. Prostate cancer is the most common nonskin neoplasm and second leading cause of death in men. 6-Shogaol (6-SHO), a potent bioactive compound in ginger (Zingiber officinale Roscoe), has been shown to possess anti-inflammatory and anticancer activity. In the present study, the effect of 6-SHO on the growth of prostate cancer cells was investigated. 6-SHO effectively reduced survival and induced apoptosis of cultured human (LNCaP, DU145, and PC3) and mouse (HMVP2) prostate cancer cells. Mechanistic studies revealed that 6-SHO reduced constitutive and interleukin (IL)-6-induced STAT3 activation and inhibited both constitutive and TNF-α-induced NF-κB activity in these cells. In addition, 6-SHO decreased the level of several STAT3 and NF-κB-regulated target genes at the protein level, including cyclin D1, survivin, and cMyc and modulated mRNA levels of chemokine, cytokine, cell cycle, and apoptosis regulatory genes (IL-7, CCL5, BAX, BCL2, p21, and p27). 6-SHO was more effective than two other compounds found in ginger, 6-gingerol, and 6-paradol at reducing survival of prostate cancer cells and reducing STAT3 and NF-κB signaling. 6-SHO also showed significant tumor growth inhibitory activity in an allograft model using HMVP2 cells. Overall, the current results suggest that 6-SHO may have potential as a chemopreventive and/or therapeutic agent for prostate cancer and that further study of this compound is warranted., (©2014 American Association for Cancer Research.)

In the present study, the ability of metformin to inhibit skin tumor promotion by 12-O-tetradecanoylphorbol-13-acetate (TPA) was analyzed in mice maintained on either an overweight control diet or an obesity-inducing diet. Rapamycin was included for comparison, and a combination of metformin and rapamycin was also evaluated. Metformin (given in the drinking water) and rapamycin (given topically) inhibited development of both papillomas and squamous cell carcinomas in overweight and obese mice in a dose-dependent manner. A low-dose combination of these two compounds displayed an additive inhibitory effect on tumor development. Metformin treatment also reduced the size of papillomas. Interestingly, all treatments seemed to be at least as effective for inhibiting tumor formation in obese mice, and both metformin and rapamycin were more effective at reducing tumor size in obese mice compared with overweight control mice. The effect of metformin on skin tumor development was associated with a significant reduction in TPA-induced epidermal hyperproliferation. Furthermore, treatment with metformin led to activation of epidermal AMP-activated protein kinase (AMPK) and attenuated signaling through mTOR complex (mTORC)-1 and p70S6K. Combinations of metformin and rapamycin were more effective at blocking epidermal mTORC1 signaling induced by TPA consistent with the greater inhibitory effect on skin tumor promotion. Collectively, the current data demonstrate that metformin given in the drinking water effectively inhibited skin tumor promotion in both overweight and obese mice and that the mechanism involves activation of epidermal AMPK and attenuated signaling downstream of mTORC1., (©2013 AACR.)